Effective tissue repair relies on an organs ability to control the number and function of stem cells or tissue specific progenitors. Satellite cells (SCs) are the "cell of choice" for adult skeletal muscle repair. Based on the assumption that all somatic cells, including SCs, have a finite capacity;reducing the number of SCs within the pool while keeping intrinsic capacity unchanged will lead to eventual functional exhaustion and impaired muscle repair. Similarly, if intrinsic capacity is impaired, while the number of SCs in the pool is maintained, this will also lead to a detrimental outcome for muscle function. During aging, muscle regeneration capacity is significantly impaired. This coincides with a decline in SC number and function. It remains unresolved whether age related impairment in muscle regeneration is caused by a loss in SC number and/or function. The first part of the proposal focuses on understanding the heterogeneity within the SC pool throughout life, moreover whether subsets of SCs are 'age-resistant. We will use a novel GFP reporter that allows proliferative output to be determined on a cell-by-cell basis in vivo. Slow and fast dividing SCs will be tested for functional diversity. This aim will answer whether aging is associated with a loss of specific functional subpopulations of SCs. The second part of the proposal focuses on studying the importance of the number of SCs available in the pool for effective repair. We will systematically decrease the number of SCs within adult and aged muscle to ask, 1) whether a decline in SC number causes impaired regeneration, 2) whether forced proliferative demand induced by limiting the number of SCs forces premature SC exhaustion. This aim will be achieved using novel genetic strategies that enable SC specific cell ablation. The third part of the proposal focuses on the intrinsic capacity of SCs to effectively repair muscle, in particular the role of P16INK4A as a regulator of SC progenitor proliferation and lineage progression during aging. P16INK4A, an aging biomarker, has been implicated in regenerative impairment of aged tissue. The role of P16INK4A in myogenesis has not been studied previously. Finally by combining SC-ablation technology and P16INK4A loss-of-function approaches, the interaction between SC number and function for effective tissue repair will be interrogated. Understanding the coordination between the number and intrinsic capacity of SCs will be critical for treatment of sarcopenia and other muscle degeneration pathologies. The specific aims of this proposal are: 1) to study heterogeneity in the SC pool as it relates to proliferative output throughout life, 2) to determine if proliferative output is causally related to sarcopenia and 3) to study the effect of genetically manipulating SC function through P16INK4A loss-of-function approaches for efficient muscle repair in young and aged muscle. PUBLIC HEALTH RELEVANCE: Deciphering the coordination between satellite cell number and function during muscle regeneration is critical for harnessing their potential to treat sarcopenia in an increasingly aging population. Furthermore, the biology of adult satellite cells may serve as a paradigm for stem cells in other tissues.